The Development and Function of Regulatory B Cells Expressing IL-10 (B10 Cells) Requires Antigen Receptor Diversity and TLR Signals

Abstract

Autoimmunity and inflammation are controlled in part by regulatory B cells, including a recently identified IL-10-competent CD1dhiCD5+ B cell subset termed B10 cells that represents 1–3% of adult mouse spleen B cells. In this study, pathways that influence B10 cell generation and IL-10 production were identified and compared with previously described regulatory B cells. IL-10-competent B cells were predominantly CD1dhiCD5+ in adult spleen and were the prevalent source of IL-10 but not other cytokines. B10 cell development and/or maturation in vivo required Ag receptor diversity and intact signaling pathways, but not T cells, gut-associated flora, or environmental pathogens. Spleen B10 cell frequencies were significantly expanded in aged mice and mice predisposed to autoimmunity, but were significantly decreased in mouse strains that are susceptible to exogenous autoantigen-induced autoimmunity. LPS, PMA, plus ionomycin stimulation in vitro for 5 h induced B10 cells to express cytoplasmic IL-10. However, prolonged LPS or CD40 stimulation (48 h) induced additional adult spleen CD1dhiCD5+ B cells to express IL-10 following PMA+ionomycin stimulation. Prolonged LPS or CD40 stimulation of newborn spleen and adult blood or lymph node CD1dlo and/or CD5− B cells also induced cytoplasmic IL-10 competence in rare B cells, with CD40 ligation uniformly inducing CD5 expression. IL-10 secretion was induced by LPS signaling through MyD88-dependent pathways, but not following CD40 ligation. LPS stimulation also induced rapid B10 cell clonal expansion when compared with other spleen B cells. Thereby, both adaptive and innate signals regulate B10 cell development, maturation, CD5 expression, and competence for IL-10 production.

IL-10-producing regulatory B cells (B10 cells) in autoimmune disease

“…B10 cell regulation of inflammation and autoimmunity is Ag specific [23,30,36]. The importance of BCR diversity is demonstrated by the fact that B10+ B10 PRO cells are reduced by approximately 90% in transgenic mice with a fixed BCR [37]. Signaling through the BCR appears critical during early development in vivo .…”

“…The combination of anti-IgM stimulation with CD40 ligation and LPS or CpG significantly reduces IL-10 competence [37]. BCR-generated signals thus inhibit the abilities of LPS or CpG and CD40 ligation to induce cytoplasmic IL-10 production.…”

“…Nevertheless, IL-21R, MHC-II and CD40 appear to be required for B10 cell effector functions, at least in experimental autoimmune encephalomyelitis (EAE) [36]. Regulatory B10 cell function therefore requires IL-21R signaling, as well as CD40 and MHC-II interactions, potentially explaining Ag-specific B10 cell effector function [37]. …”

“…B10 cells are present in T cell-deficient nude mice, and their frequencies and numbers are approximately fivefold higher when compared with wildtype mice. This observation is strengthened by the fact that MHC class I and MHC-II molecules and CD1d expression are not required for B10 cell development [37]. The presence or absence of T cells in vitro also does not affect the frequency of B10 cells.…”

“…Mouse B10 PRO cells acquire the ability to function like B10 cells after in vitro maturation following stimulation with LPS, but not CpG, in the presence or absence of agonistic CD40 mAb [32]. TLR4 and TLR9 signaling through myeloid differentiation primary response gene 88 (MyD88) is necessary for the optimal maturation and IL-10 induction of B10pro and B10 cells following LPS stimulation and LPS or CpG stimulation, respectively [37]. Nevertheless, MyD88 expression is not an absolute requirement for B10 cell development in vivo , since B10 cells develop normally in MyD88 --/-- mice [37].…”